Prenylation, the attachment of a farnesyl or geranylgeranyl isoprenoid to a C-terminal cysteine, is a post-translational modification which modulates localisation and function of key proteins such as members of the RAS superfamily, nuclear lamins and heterotrimeric G-proteins. In humans, three enzymes catalyse the prenylation reaction: farnesyl transferase, geranylgeranyl transferase type 1 and RAB geranylgeranyl transferase.
Substantial effort has gone into developing therapeutics that disrupt prenylation as a means of targeting RAS-driven cancers, yet these approaches have failed to deliver in clinical trials. This is largely due to the fact that the dynamics of prenylation in response to prenyl transferase inhibitors is more complex than originally envisaged and includes changes in prenylation state and altered substrate recognition by the different transferase enzymes. Emerging research shows that prenylation also plays a role in cardiovascular and neurodegenerative diseases, Hutchinson-Gilford progeria syndrome (HGPS), choroideremia and viral infections.
Advances in the study of protein prenylation are hampered by the lack of inherent handles that allow for isolation and identification of the prenylated proteome. Novel approaches for profiling the prenylated proteome, its dynamics, and effects of varied inhibitors of the prenyl transferases are imperative for explaining past failures and to direct future studies.
This thesis presents a novel set of isoprenoid analogues that can selectively label prenylated proteins in cells. The application of these probes in combination with quantitative proteomic techniques enabled the description of the complex dynamics between different isoprenoid substrates and transferases in response to prenyl transferase inhibitors, and to validate several novel prenylated substrates. We describe attempts to profile prenylation in cells derived from HGPS patients, and explore potential targets of farnesyl transferase inhibition in pulmonary arterial cells. We envisage that the robust methodology presented herein will be widely applicable to the study of prenylation in the context of both health and disease.